It has been postulated that the determinants of neuronal cell death in acute and chronic neurodegenerative conditions are mediated by free radical damage. Epicatechin and cocoa (Theobroma cacao), has been reported to be neuroprotective and a potential preventive medicine, but the underlying cellular mechanisms are still unclear. Our preliminary results prompted us to focus our attention on (-)-epicatechin and flavanol-rich cocoa extract and test the hypothesis that heme oxygenase (HO) activity could participate in cocoa extract neuroprotective function. HO, which cleaves heme (a prooxidant) to form biliverdin/bilirubin (antioxidants), carbon monoxide (a vasodilator), and iron (which by increasing ferritin would be protective) has been shown to play a protective role in oxidative stress, ischemia, inflammation, and hypertension. Although HO2 is constitutively expressed, HO1 is inducible. Consequently, an increase in the activity of this endogenous antioxidant system via an increase in HO1 levels, could be a way to achieve neuroprotection both at a cellular and organ levels. Of the compounds tested in our preliminary experiments using primary neuronal cultures, epicatechin was one of the most potent HO1 inducers. Our results using stroke models also suggest that epicatechin given orally significantly prevented ischemic-reperfusion brain damage. Together, our results indicate that pretreatment with the single compound epicatechin which is the flavanol most enriched in cocoa extract - and likely to cocoa extract itself - is sufficient to provide endogenous neuroprotection, suggesting that co-treatment during oxidative stress is not necessary. These preliminary results implied that specific induction of HO1 could be a mechanism by which cocoa extract exerts its neuroprotective actions and motivated us to propose that some of the neuroprotective effects attributed to Cocoa extract could be mediated through a pathway leading to stimulation of an endogenous antioxidant pathway. In Aim 1, we will determine neuronal cell death and behavioral outcomes following global ischemia in wildtype (WT) mice pre-treated (acutely or chronically) with epicatechin and/or flavanol-rich cocoa extract and test whether these effects are attenuated in knockout mice. In Aim 2, we will determine whether changes in HO1 expression induced by epicatechin and/or flavanol-rich cocoa extract result in changes in cell survival in neuronal cultures derived from WT and knockout mice. Together, these results will help us determine whether consumption of a standardized cocoa extract could be beneficial and the pathways by which cocoa extract could provide the brain with resistance to acute debilitating neurodegenerative conditions. PUBLIC HEALTH RELEVANCE: For centuries, cocoa (Theobroma cacao) has been reported as preventive medicine to strengthen the nervous system, but the underlying cellular mechanisms are still unclear. Our preliminary results prompted us to focus our attention on epicatechin and cocoa extract and test the hypothesis that the pathway leading to heme oxygenase (HO) enzyme induction could participate in cocoa's neuroprotective function. HO, which cleaves heme (a pro-oxidant) to form biliverdin/bilirubin (anti-oxidants) has been shown to play a protective role in oxidative stress, ischemia, inflammation, and hypertension. Using pre-clinical laboratory mouse models, we will determine whether prophylactic consumption of a standardized flavanol-rich cocoa extract can prevent neurological decline and neuronal cell death following global ischemia;thus providing new pathways by which cocoa could provide brain resistance against acute neurological disorders.